PROJECT SUMMARY/ABSTRACT Rotator cuff tears are common, disabling, and are costly musculoskeletal injuries. Surgical repair is a common treatment, but recurrent tears occur in 20 to 90% of patients, especially in those with large or massive tears. Rotator cuff repair with biomaterial augmentations, including mechanical augmentation to increase the repair strength and biological healing to accelerate tissue regeneration, may reduce postoperative complications. However, the current graft materials used for augmentation are far from satisfactory. Tissue engineering offers the potential to generate functional tissue replacement, however, designing an appropriate scaffold that can has native tendon mechanical properties and provide an optimal environment for cell seeding, growth, and differentiation, especially for tendon enthesis regeneration, has proven challenging. In this application, we propose a novel tissue engineering approach using a sliced tendon fibrocartilage bone composite (TFBC) as a scaffold to seed bone marrow-derived stem cells (BMSC) for rotator cuff repair augmentation. Our preliminary studies indicated that this TFBC can mechanically enhance the rotator cuff repair. The TFBC can be revitalized by seeding BMSCs, which express tenogenesis in native tendon environment under mechanical loading. Our recent in vivo data demonstrated that our engineered TFBC biologically augmented the rotator cuff repair better when compared to repair or TFBC scaffold alone after six weeks post-surgery in our newly developed canine non-weight bearing shoulder model which mimics human shoulder function and activities. Based on our preliminary studies, the overall goal of the current proposal is to develop and test a functional engineered composite tissue (TFBC) for rotator tear treatment. Our underlying hypothesis is that our engineered TFBC, as an augmentation biomaterial, will improve functional outcomes following rotator cuff repairs compared to the clinically used biomaterial (GraftJacket). We also postulate that TFBC will help to rebuild the tendon enthesis, which has been a great challenge to regenerate in the current approaches, as the TFBC contains a native fibrocartilage zone. To test our hypothesis, the following two specific aims are proposed. Specific Aim 1 is to test the TFBC mechanical augmentation properties when the TFBC is used for rotator cuff repair in an in vitro model. Specific Aim 2 is to test engineered TFBC biological augmentation and enthesis regeneration in a canine in vivo model with a long-term followup. We expect two important outcomes from this investigation: 1) The TFBC strengthens the mechanical properties of the rotator cuff repair, and 2) the engineered TFBC effectively reduce the rotator repair failure, improve the rotator cuff repair healing, and rebuild a tendon enthesis which has not been successfully regenerate to date. If our hypothesis is supported, we would have developed a novel, effective, and clinically-applicable biomaterial to improve the quality of rotator cuff repairs, which has a significant impact on the field of rotator cuff tear, a challenging clinical and research issue.